PROJECT SUMMARY Alphaviruses are positive-strand enveloped RNA viruses of the Togaviridae family that globally cause disease in humans. As an example, Chikungunya virus (CHIKV) is a mosquito-transmitted alphavirus that causes explosive epidemics in humans of a severe febrile illness characterized by debilitating acute and chronic polyarthritis. Other arthritogenic alphaviruses (e.g., Ross River (RRV), Mayaro (MAYV), and O?nyong nyong (ONNV) viruses) are emerging beyond their historical boundaries with outbreaks in Oceania, Africa, and the Americas. Venezuelan (VEEV) and Eastern (EEEV) equine encephalitis viruses are mosquito-transmitted alphaviruses with the potential to cause fatal neuroinvasive disease in humans. At present, no antiviral agents or licensed vaccines exist for the treatment or prevention of any alphavirus infections. The goal of this highly interactive Project 3 between long-standing collaborators (Diamond, Crowe, Kuhn/Rossmann, Fremont, and Streblow laboratories) and a new commercial partner (IDBiologics) is to develop Fc-optimized, high potency pan-alphavirus human mAbs for the prevention and treatment of multiple alphavirus infections in humans. Our approach is to screen existing and newly generated panels of human mAbs derived from subjects who were infected or immunized previously with different alphaviruses (CHIKV, RRV, or VEEV) for extensive cross-reactivity, neutralization, and binding to the surface of infected cells. After a stringent filtering and Go/No-Go decision process, lead candidate mAbs will be improved by affinity maturation and optimization of Fc effector functions, and therapeutic potential will be tested in murine models of alphavirus-induced arthritis and encephalitis. Two secondary goals are to define the structural and mechanistic correlates of human mAb protection. This information will inform a possible combination mAb strategy. Subsequently, we will generate CHO cell lines for IND enabling studies, and pairs of optimized pan-alphavirus mAbs will be tested for therapeutic activity in non-human primate (NHP) models of alphavirus infection. The proposed research achieves the following CETR RFA goals: (a) Basic discoveries that are translated into treatments by generating pan-virus mAb therapy with potent protective efficacy. (b) Since many alphaviruses are emerging and cause large-scale epidemics, this project will develop a therapy for diseases of unmet clinical need. (c) The work requires an interdisciplinary academic team and industry partner, which should facilitate the translation into therapeutics. (d) Studies in this project and CETR provide a platform for the discovery of mAb therapeutics against other viruses. A pan- alphavirus mAb therapy against the majority of globally relevant alphaviruses will provide an immediate countermeasure against disease emergence or bioterrorist introduction.